Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Pseudomonas aeruginosa defends against phages through type IV pilus glycosylation


Since phages present a major challenge to survival in most environments, bacteria express a battery of anti-phage defences including CRISPR–Cas, restriction-modification and abortive infection systems1,2,3,4. Such strategies are effective, but the phage genome—which encodes potentially inhibitory gene products—is still allowed to enter the cell. The safest way to preclude phage infection is to block initial phage adsorption to the cell. Here, we describe a cell-surface modification that blocks infection by certain phages. Strains of the opportunistic pathogen Pseudomonas aeruginosa express one of five different type IV pilins (T4P)5, two of which are glycosylated with O-antigen units6 or polymers of d-arabinofuranose7,8,9. We propose that predation by bacteriophages that use T4P as receptors selects for strains that mask potential phage binding sites using glycosylation. Here, we show that both modifications protect P. aeruginosa from certain pilus-specific phages. Alterations to pilin sequence can also block phage infection, but glycosylation is considered less likely to create disadvantageous phenotypes. Through construction of chimeric phages, we show that specific phage tail proteins allow for infection of strains with glycosylated pili. These studies provide insight into first-line bacterial defences against predation and ways in which phages circumvent them, and provide a rationale for the prevalence of pilus glycosylation in nature.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Fig. 1: Infection of a P. aeruginosa PAO1 pilA mutant expressing non-glycosylated versus glycosylated pilins.
Fig. 2: A chimeric phage expressing DMS3 genes in the JBD26 background gains the ability to infect a strain with glycosylated pilins.
Fig. 3: Pilin D-region sequence affects phage susceptibility.
Fig. 4: Pilin d-arabinofuranosylation blocks phage infection.


  1. 1.

    Weinbauer, M. G. Ecology of prokaryotic viruses. FEMS Microbiol. Rev. 28, 127–181 (2004).

    CAS  Article  Google Scholar 

  2. 2.

    Sampson, T. R. & Weiss, D. S. Alternative roles for CRISPR/Cas systems in bacterial pathogenesis. PLoS Pathog. 9, e1003621 (2013).

    Article  Google Scholar 

  3. 3.

    Sorek, R., Lawrence, C. M. & Wiedenheft, B. CRISPR-mediated adaptive immune systems in bacteria and archaea. Annu. Rev. Biochem. 82, 237–266 (2013).

    CAS  Article  Google Scholar 

  4. 4.

    Labrie, S. J., Samson, J. E. & Moineau, S. Bacteriophage resistance mechanisms. Nat. Rev. Microbiol. 8, 317–327 (2010).

    CAS  Article  Google Scholar 

  5. 5.

    Kus, J. V., Tullis, E., Cvitkovitch, D. G. & Burrows, L. L. Significant differences in type IV pilin allele distribution among Pseudomonas aeruginosa isolates from cystic fibrosis (CF) versus non-CF patients. Microbiology 150, 1315–1326 (2004).

    CAS  Article  Google Scholar 

  6. 6.

    DiGiandomenico, A. et al. Glycosylation of Pseudomonas aeruginosa 1244 pilin: glycan substrate specificity. Mol. Microbiol. 46, 519–530 (2002).

    CAS  Article  Google Scholar 

  7. 7.

    Kus, J. V. et al. Modification of Pseudomonas aeruginosa Pa5196 type IV pilins at multiple sites with d-Araf by a novel GT-C family Arabinosyltransferase, TfpW. J. Bacteriol. 190, 7464–7478 (2008).

    CAS  Article  Google Scholar 

  8. 8.

    Harvey, H., Kus, J. V., Tessier, L., Kelly, J. & Burrows, L. L. Pseudomonas aeruginosa d-arabinofuranose biosynthetic pathway and its role in type IV pilus assembly. J. Biol. Chem. 286, 28128–28137 (2011).

    CAS  Article  Google Scholar 

  9. 9.

    Voisin, S. et al. Glycosylation of Pseudomonas aeruginosa strain Pa5196 type IV pilins with mycobacterium-like α-1,5-linked d-Araf oligosaccharides. J. Bacteriol. 189, 151–159 (2007).

    CAS  Article  Google Scholar 

  10. 10.

    Lam, J. S., Taylor, V. L., Islam, S. T., Hao, Y. & Kocincova, D. Genetic and functional diversity of Pseudomonas aeruginosa lipopolysaccharide. Front. Microbiol. 2, 118 (2011).

    CAS  Article  Google Scholar 

  11. 11.

    Bradley, D. E. & Pitt, T. L. Pilus-dependence of four Pseudomonas aeruginosa bacteriophages with non-contractile tails. J. Gen. Virol. 24, 1–15 (1974).

    CAS  Article  Google Scholar 

  12. 12.

    Burrows, L. L. Pseudomonas aeruginosa twitching motility: type IV pili in action. Annu. Rev. Microbiol. 66, 493–520 (2012).

    CAS  Article  Google Scholar 

  13. 13.

    Craig, L. & Li, J. Type IV pili: paradoxes in form and function. Curr. Opin. Struct. Biol. 18, 267–277 (2008).

    CAS  Article  Google Scholar 

  14. 14.

    Heiniger, R. W., Winther-Larsen, H. C., Pickles, R. J., Koomey, M. & Wolfgang, M. C. Infection of human mucosal tissue by Pseudomonas aeruginosa requires sequential and mutually dependent virulence factors and a novel pilus-associated adhesin. Cell. Microbiol. 12, 1158–1173 (2010).

    CAS  Article  Google Scholar 

  15. 15.

    Ohama, M. et al. Intratracheal immunization with pili protein protects against mortality associated with Pseudomonas aeruginosa pneumonia in mice. FEMS Immunol. Med. Microbiol. 47, 107–115 (2006).

    CAS  Article  Google Scholar 

  16. 16.

    Hahn, H. P. The type-4 pilus is the major virulence-associated adhesin of Pseudomonas aeruginosa – a review. Gene 192, 99–108 (1997).

    CAS  Article  Google Scholar 

  17. 17.

    Bucior, I., Pielage, J. F. & Engel, J. N. Pseudomonas aeruginosa pili and flagella mediate distinct binding and signaling events at the apical and basolateral surface of airway epithelium. PLoS Pathog. 8, e1002616 (2012).

    CAS  Article  Google Scholar 

  18. 18.

    James, C. E. et al. Differential infection properties of three inducible prophages from an epidemic strain of Pseudomonas aeruginosa. BMC Microbiol. 12, 216 (2012).

    CAS  Article  Google Scholar 

  19. 19.

    Bondy-Denomy, J. et al. Prophages mediate defense against phage infection through diverse mechanisms. ISME J. 10, 2854–2866 (2016).

    Article  Google Scholar 

  20. 20.

    Nguyen, Y. et al. Pseudomonas aeruginosa minor pilins prime type IVa pilus assembly and promote surface display of the PilY1 adhesin. J. Biol. Chem. 290, 601–611 (2015).

    CAS  Article  Google Scholar 

  21. 21.

    Winther-Larsen, H. C. et al. Pseudomonas aeruginosa type IV pilus expression in Neisseria gonorrhoeae: effects of pilin subunit composition on function and organelle dynamics. J. Bacteriol. 189, 6676–6685 (2007).

    CAS  Article  Google Scholar 

  22. 22.

    Giltner, C. L., Habash, M. & Burrows, L. L. Pseudomonas aeruginosa minor pilins are incorporated into type IV pili. J. Mol. Biol. 398, 444–461 (2010).

    CAS  Article  Google Scholar 

  23. 23.

    Comer, J. E., Marshall, M. A., Blanch, V. J., Deal, C. D. & Castric, P. Identification of the Pseudomonas aeruginosa 1244 pilin glycosylation site. Infect. Immun. 70, 2837–2845 (2002).

    CAS  Article  Google Scholar 

  24. 24.

    Smedley, J. G. III et al. Influence of pilin glycosylation on Pseudomonas aeruginosa 1244 pilus function. Infect. Immun. 73, 7922–7931 (2005).

    CAS  Article  Google Scholar 

  25. 25.

    Asikyan, M. L., Kus, J. V. & Burrows, L. L. Novel proteins that modulate type IV pilus retraction dynamics in Pseudomonas aeruginosa. J. Bacteriol. 190, 7022–7034 (2008).

    CAS  Article  Google Scholar 

  26. 26.

    Allison, T. M., Conrad, S. & Castric, P. The group I pilin glycan affects type IVa pilus hydrophobicity and twitching motility in Pseudomonas aeruginosa 1244. Microbiology 161, 1780–1789 (2015).

    CAS  Article  Google Scholar 

  27. 27.

    Gault, J. et al. Neisseria meningitidis type IV pili composed of sequence invariable pilins are masked by multisite glycosylation. PLoS Pathog. 11, e1005162 (2015).

    Article  Google Scholar 

  28. 28.

    Piepenbrink, K. H. et al. Structural diversity in the type IV pili of multidrug-resistant Acinetobacter. J. Biol. Chem. 291, 22924–22935 (2016).

    CAS  Article  Google Scholar 

  29. 29.

    Harvey, H., Habash, M., Aidoo, F. & Burrows, L. L. Single-residue changes in the C-terminal disulfide-bonded loop of the Pseudomonas aeruginosa type IV pilin influence pilus assembly and twitching motility. J. Bacteriol. 191, 6513–6524 (2009).

    CAS  Article  Google Scholar 

  30. 30.

    Heo, Y. J., Chung, I. Y., Choi, K. B., Lau, G. W. & Cho, Y. H. Genome sequence comparison and superinfection between two related Pseudomonas aeruginosa phages, D3112 and MP22. Microbiology 153, 2885–2895 (2007).

    CAS  Article  Google Scholar 

  31. 31.

    Castric, P., Cassels, F. J. & Carlson, R. W. Structural characterization of the Pseudomonas aeruginosa 1244 pilin glycan. J. Biol. Chem. 276, 26479–26485 (2001).

    CAS  Article  Google Scholar 

  32. 32.

    Tan, R. M. et al. Type IV pilus glycosylation mediates resistance of Pseudomonas aeruginosa to opsonic activities of the pulmonary surfactant protein A. Infect. Immun. 83, 1339–1346 (2015).

    CAS  Article  Google Scholar 

  33. 33.

    Nguyen, Y. et al. Structural and functional studies of the Pseudomonas aeruginosa minor pilin, PilE. J. Biol. Chem. 290, 26856–26865 (2015).

    CAS  Article  Google Scholar 

  34. 34.

    Craig, L. et al. Type IV pilin structure and assembly: X-ray and EM analyses of Vibrio cholerae toxin-coregulated pilus and Pseudomonas aeruginosa PAK pilin. Mol. Cell. 11, 1139–1150 (2003).

    CAS  Article  Google Scholar 

  35. 35.

    Kolappan, S. et al. Structure of the Neisseria meningitidis type IV pilus. Nat. Commun. 7, 13015 (2016).

    CAS  Article  Google Scholar 

  36. 36.

    Davidson, A. R., Cardarelli, L., Pell, L. G., Radford, D. R. & Maxwell, K. L. Long noncontractile tail machines of bacteriophages. Adv. Exp. Med. Biol. 726, 115–142 (2012).

    CAS  Article  Google Scholar 

  37. 37.

    Le, S. et al. Mapping the tail fiber as the receptor binding protein responsible for differential host specificity of Pseudomonas aeruginosa bacteriophages PaP1 and JG004. PLoS ONE 8, e68562 (2013).

    CAS  Article  Google Scholar 

  38. 38.

    Bondy-Denomy, J., Pawluk, A., Maxwell, K. L. & Davidson, A. R. Bacteriophage genes that inactivate the CRISPR/Cas bacterial immune system. Nature 493, 429–432 (2013).

    CAS  Article  Google Scholar 

  39. 39.

    Schneider, C. A., Rasband, W. S. & Eliceiri, K. W. NIH Image to ImageJ: 25 years of image analysis. Nat. Methods 9, 671–675 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Hoang, T. T., Karkhoff-Schweizer, R. R., Kutchma, A. J. & Schweizer, H. P. A broad-host-range Flp-FRT recombination system for site-specific excision of chromosomally-located DNA sequences: application for isolation of unmarked Pseudomonas aeruginosa mutants. Gene 212, 77–86 (1998).

    CAS  Article  Google Scholar 

  41. 41.

    Lavigne, R., Ceyssens, P. J. & Robben, J. Phage proteomics: applications of mass spectrometry. Methods Mol. Biol. 502, 239–251 (2009).

    CAS  Article  Google Scholar 

Download references


We thank K. Maxwell for helpful comments on the manuscript. This work was supported by Canadian Institutes of Health Research Open Operating Grants to L.L.B. (MOP 86639) and to A.R.D. (XNE-86943 and MOP-115039).

Author information




H.H., A.R.D. and L.L.B. designed the study; H.H., J.B.-D., H.M. and K.M.S. performed experiments; H.H., A.R.D. and L.L.B. analysed the data; A.R.D. and L.L.B. wrote the manuscript with input from H.H., J.B.-D. and H.M. All authors approved the final version.

Corresponding authors

Correspondence to Alan R. Davidson or Lori L. Burrows.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Supplementary Information

Supplementary Figures 1–3, Supplementary Figure References, Supplementary Tables 1–5, Supplementary Table 5 References.

Life Sciences Reporting Summary

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Harvey, H., Bondy-Denomy, J., Marquis, H. et al. Pseudomonas aeruginosa defends against phages through type IV pilus glycosylation. Nat Microbiol 3, 47–52 (2018).

Download citation

Further reading


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing